Thymidine kinase expressing natural killer cell lines and methods of use

Abstract

This invention relates to a natural killer cell line termed NK-92. The invention provides a vector for transfecting a mammalian cell which includes a nucleic acid sequence encoding a cytokine that promotes the growth of NK-92. Additionally, the invention provides an NK-92 cell, or an NK-92 cell modified by transfection with a vector conferring advantageous properties, which is unable to proliferate and which preserves effective cytotoxic activity. The invention further provides a modified NK-92 cell that is transfected with a vector encoding a cytokine that promotes the growth of NK-92 cells. The cell secretes the cytokine upon being cultured under conditions that promote cytokine secretion, and furthermore secretes the cytokine in vivo upon being introduced into a mammal. In a significant embodiment, the cytokine is interleukin 2. The present invention also provides methods of purging cancer cells from a biological sample, of treating a cancer ex vivo in a mammal, and of treating a cancer in vivo in a mammal employing a natural killer cell, such as NK-92 itself, an NK-92 cell which is unable to proliferate and which preserves effective cytotoxic activity, or natural killer cells transfected with a vector encoding a cytokine.

That which we claim is:  
     
         1 . A method of purging cells related to a pathology from a biological sample, said method comprising (i) obtaining a biological sample from a mammal, wherein the biological sample is suspected of containing cells related to the pathology; and (ii) contacting the biological sample with a medium comprising NK-92 or modified NK-92 natural killer cells, wherein the modified NK-92 cells have been modified by a physical treatment or by transfection with a vector; whereby the natural killer cells purge cells related to the pathology from the sample.  
     
     
         2 . The method described in  claim 1  wherein the pathology is a cancer.  
     
     
         3 . The method described in  claim 1  wherein the pathology is an infection by a pathogenic virus.  
     
     
         4 . The method described in  claim 3  wherein the pathogenic virus is human immunodeficiency virus, Epstein-Barr virus, cytomegalovirus, or herpes virus.  
     
     
         5 . The method described in  claim 1  wherein the biological sample is human blood or bone marrow.  
     
     
         6 . The method described in  claim 1  wherein the natural killer cell is immobilized on a support.  
     
     
         7 . The method described in  claim 1  wherein the modified NK-92 cells have been modified by a physical treatment that renders them non-proliferative, said treatment not significantly diminishing their cytotoxicity, by treatment that inhibits expression of HLA antigens on the NK-92 cell surface, by transfection with a vector, or by any combination thereof.  
     
     
         8 . The method described in  claim 7  wherein the cells have been transfected with a vector encoding a cytokine that promotes the growth of the cells, a vector encoding a protein that is responsive to an agent, a vector encoding a cancer cell receptor molecule, or with any combination thereof.  
     
     
         9 . The method described in  claim 1  wherein the medium further comprises cytokine that promotes the growth of the cells.  
     
     
         10 . A method of treating a pathology ex vivo in a mammal comprising the steps of: 
 (i) obtaining a biological sample from the mammal, wherein the sample is suspected of containing cells related to the pathology;    (ii) contacting the biological sample with a medium comprising NK-92 or modified NK-92 natural killer cells, wherein the modified NK-92 cells have been modified by a physical treatment or by transfection with a vector, whereby the cells related to the pathology in the sample are selectively destroyed, thereby producing a purged sample; and    (iii) returning the purged sample to the mammal.    
     
     
         11 . The method described in  claim 10  wherein the pathology is a cancer.  
     
     
         12 . The method described in  claim 11  wherein the cancer is a leukemia, a lymphoma or a multiple myeloma.  
     
     
         13 . The method described in  claim 10  wherein the pathology is an infection by a pathogenic virus.  
     
     
         14 . The method described in  claim 13  wherein the pathogenic virus is human immunodeficiency virus, Epstein-Barr virus, cytomegalovirus, or herpes virus.  
     
     
         15 . The method described in  claim 10  wherein the biological sample is blood or bone marrow and wherein the mammal is a human.  
     
     
         16 . The method described in  claim 10  wherein the natural killer cell is immobilized on a support.  
     
     
         17 . The method described in  claim 10  wherein the medium comprises modified NK-92 cells which have been modified by a physical treatment that renders them non-proliferative, said treatment not significantly diminishing their cytotoxicity, by treatment that inhibits expression of HLA antigens on the NK-92 cell surface, by transfection with a vector, or by any combination thereof.  
     
     
         18 . The method described in  claim 17  wherein the cells have been transfected with a vector encoding a cytokine that promotes the growth of the cells, a vector encoding a protein that is responsive to an agent, a vector encoding a cancer cell receptor molecule, or with any combination thereof.  
     
     
         19 . The method of treating a cancer described in  claim 10  wherein the medium further comprises a cytokine that promotes the growth of the cells.  
     
     
         20 . A method of treating a pathology in vivo in a mammal comprising the step of administering to the mammal a medium comprising NK-92 or modified NK-92 natural killer cells, wherein the modified NK-92 cells have been modified by a physical treatment that renders them non-proliferative, said treatment not significantly diminishing their cytotoxicity, by treatment that inhibits expression of HLA antigens on the NK-92 cell surface, by transfection with a vector, or by any combination thereof.  
     
     
         21 . The method described in  claim 20  wherein the modified NK-92 cells have been transfected with a vector encoding a cytokine that promotes the growth of the cells, with a vector encoding a protein that is responsive to an agent, a vector encoding a cancer cell receptor molecule, or with any combination thereof.  
     
     
         22 . The method described in  claim 20  wherein the pathology is a cancer.  
     
     
         23 . The method of treating a cancer described in  claim 22  wherein the cancer is a leukemia, a lymphoma or a multiple myeloma.  
     
     
         24 . The method described in  claim 20  wherein the pathology is an infection by a pathogenic virus.  
     
     
         25 . The method described in  claim 24  wherein the pathogenic virus is human immunodeficiency virus, Epstein-Barr virus, cytomegalovirus, or herpes virus.  
     
     
         26 . The method of treating a pathology described in  claim 20  wherein the route of administration of the cells to the mammal is intravenous and the mammal is a human.  
     
     
         27 . The method of treating a pathology described in  claim 20  further comprising administering a cytokine that promotes the growth of the cells to the mammal in conjunction with administering the medium comprising the natural killer cell.  
     
     
         28 . The method of treating a pathology described in  claim 26  wherein the NK-92 is modified by transfection with a vector encoding a protein that is responsive to an agent such that when the agent is taken up by the cell, the cell is inactivated, and wherein the method further comprises administering to the mammal, after a time sufficient for the natural killer cell to treat the cancer has elapsed, an amount of the agent effective to inactivate the cell.  
     
     
         29 . The method of treating a pathology described in  claim 28  wherein the agent is acyclovir or gancyclovir.